Loss of replicative ability, which may represent "aging" will be examined in human diploid fibroblasts. Since continued DNA synthesis is required for the survival of dividing cells and since damage to DNA or to the replication process may be involved in aging, a search will be made for age- or passage-associated changes in the orderly process of DNA synthesis. DNA synthesis will be examined in both intact and permeabilized cells (cells rendered permeable to various metabolites) by determining the rate and extent of incorporation of labeled precursors at the cellular level and by analyzing replicon synthesis from DNA fiber-autoradiograms prepared from such cells. Any change in DNA synthesis observed at the cellular level or any change observed at the replicon level will be investigated, in detail, to determine if it is due to an alteration in the DNA template, an alteration in the DNA polymerases, or an alteration in the amount of metabolites present inside cells. The proposed experiments will involve mixing cells, or extracts of cells, and examining DNA synthesis under permeable conditions. Finally, the repair potential of cells of different "ages" will be determined by examining the short term and long term effects of ionizing radiation and ultraviolet (UV) radiation on DNA replication patterns.